The present invention relates to methods of treating isolated pancreatic cells and microencapsulated pancreatic cells, in order to prepare the cells for transplantation. Media containing an antibiotic, an anti-oxidant, an anti-cytokine, or an anti-endotoxin, or combinations thereof, are utilized.
Glycemic control in diabetes has been shown to delay the onset of, and slow the progression of, associated pathological complications. However, achieving adequate glycemic control using insulin therapy can be difficult. One alternative to insulin therapy is the transplantation of functioning pancreatic islet cells to diabetic subjects, to provide biological insulin replacement.
Approximately one percent of the volume of the human pancreas is made up of islets of Langerhans (hereinafter xe2x80x9cisletsxe2x80x9d), which are scattered throughout the exocrine pancreas. Each islet comprises insulin producing beta cells as well as glucagon containing alpha cells, somatostatin secreting delta cells, and pancreatic polypeptide containing cells (PP-cells). The majority of islet cells are insulin-producing beta cells.
However, transplanted or grafted islet cells encounter immunological rejection, which can limit the clinical usefulness of this method. (Brunicardi and Mullen, Pancreas 9:281 (1994); Bartlett et al., Transplant. Proc. 26:737 (1994)). To combat rejection, immunosuppressive drugs may be used, but such immunosuppressive therapy impairs the body""s immunological defenses and carries significant side effects and risks in itself. Approaches to containing and protecting transplanted islet cells have been proposed, including the use of extravascular diffusion chambers, intravascular diffusion chambers, intravascular ultrafiltration chambers, macroencapsulation and microencapsulation. The goal of islet transplantation is to achieve normoglycemia in the treated subject for some extended period of time.
Microencapsulation of islet cells has been proposed to reduce or avoid immunological rejection of transplanted islet cells. Lim and Sun, Science 210:908 (1980). The cells are encapsulated in a membrane that is permeable to cell substrates and cell secretions, but essentially impermeable to bacteria, lymphocytes, and large immunological proteins. The method of microencapsulation described by Lim and Sun involves forming gelled alginate droplets around isolated islet cells, and then adding coats of poly-L-lysine and additional alginate. The inner gelled core of the microcapsule is then liquefied by chelation. However, chelation of the core affects the structural support of the capsules and may adversely affect durability. The success of microencapsulated islet cell transplantation in treating diabetes depends on the ability of the microcapsules to provide sufficient amounts of insulin in response to glucose stimulation, over an extended period of time, to achieve adequate glycemic control.
Methods of treating isolated pancreatic cells, or of treating microencapsulated pancreatic cells, to enhance glucose-stimulated insulin production by the microcapsules and to provide durable microcapsules capable of glucose-stimulated insulin production, are therefore desirable.
A first aspect of the present invention is a method of treating isolated living cells, by first culturing the cells in a medium containing at least one of (or a combination of ): an antioxidant, an anti-cytokine, an anti-endotoxin, or an antibiotic. The cells are then microencapsulated in a biocompatible microcapsule that contains a hydrogel core and a semipermeable outer membrane, to provide a microcapsule containing living cells therein.
A further aspect of the present invention is a method of treating isolated living cells, by first cryopreserving the cells in a cryopreservation medium containing at least one of (or a combination of): an antioxidant, an anti-cytokine, an anti-endotoxin, or an antibiotic; then thawing the cells and encapsulating the cells in a biocompatible microcapsule having a hydrogel core and a semipermeable outer membrane.
A further aspect of the present invention is a method of treating biocompatible microcapsules containing living cells, where the microcapsule contains a hydrogel core and a semipermeable outer membrane. The microcapsules are cultured in a medium containing at least one of (or a combination of): an antioxidant, an anti-cytokine, an anti-endotoxin, or an antibiotic.
A further aspect of the present invention is a method of preparing microencapsulated cells by first culturing the cells in a cell culture medium containing at least one of (or a combination of): antioxidants, anti-cytokines, anti-endotoxins, and antibiotics. The cells are then encapsulated in a biocompatible microcapsule having a hydrogel core and a semipermeable outer membrane, where the living cells are present in the core. The microcapsules are then cultured in a medium containing at least one of (or a combination of): an antioxidant, an anti-cytokine, an anti-endotoxin, and an antibiotic.
A further aspect of the present invention is a method of preparing microencapsulated cells that includes a step of incubating the microencapsulated cells with a physiologically acceptable salt such as sodium sulfate or the like in order to produce a more durable, and therefore useful, biocompatible microcapsule.
A further aspect of the present invention is a method of isolating pancreatic islet cells in which an antioxidant is included in the digestion medium that is used to free the islet cells from pancreatic tissue.
A further aspect of the present invention is microencapsulated cell products, which may be produced by a process as described above.